Cryptosporidium parvum is a major cause of diarrhea worldwide, for which there is no reliable antiparasitic therapy. In immunocompetent individuals, C. parvum infection results in a self-limited diarrheal illness. By contrast, AIDS patients may develop chronic diarrhea, which can be fatal. Effective antiretroviral therapy can lead to resolution of AiDS-associated cryptosporidiosis, presumably due to improvement in the intestinal immune response. The long-term goals of this project are to determine the immune mechanisms involved in the control of cryptosporidiosis in healthy adults and AIDS patients on effective antiretroviral treatment. We have demonstrated that sensitized, immunocompetent volunteers expressed interferon gamma (IFNT) in response to C. parvum exposure and that IFN7 expression was associated with resistance to infection. By contrast, naive, symptomatic individuals initially expressed Interleukin 15 (IL-15), which was associated with control of oocyst excretion. Neither IL-15 nor IFN7 was detected in AIDS-associated chronic cryptosporidiosis, but expression of IL-15 and IFN7 was noted in biopsies obtained from patients responding to antiretroviral therapy. Preliminary studies demonstrated that IL-15 can activate lymphocytes to lyse infected epithelial cells. However, many questions remain. For example, what are the effector mechanisms used by IL-15 and IFNgamma, in the control phase and how are these responses coordinated? Can Thl cytokines in fact lead to resolution of cryptosporidiosis in AIDS patients in the absence of immune recovery? What is the sequence of responses in AIDS patients with immune recovery with effective anti-retroviral therapy. The specific aims of the current proposal are: 1 ) To test the hypothesis that IL-15 and IFNgamma help clear infection of epithelial cells by activation of cytolytic cells and establish the mechanisms used by the effector cells. 2) To confirm the importance of Thl cytokines in resolution of cryptosporidiosis by conducting a pilot, proof-of-concept, open-label trial of IL-12 therapy in chronic cryptosporidiosis in AIDS patients not responding to antiretroviral therapy. 3) To confirm that mechanisms used by cytolytic cells defined in aim 1 and associated cytokines, effector molecules, and chemokines are expressed in the intestines in human cryptosporidiosis using microarray analysis of intestinal biopsies obtained before and after experimental challenge of immunocompetent adults with C. parvurn oocysts. 4) To test the hypothesis that AIDS patients with cryptosporidiosis sequentially expresses innate and then Thl memory responses during immune reconstitution. These studies should identify key aspects of the human immune response needed for vaccines to prevent cryptosporidiosis and identify the host responses that can be targeted for adjunctive immunotherapy for cryptosporidiosis in patients with AIDS and other immunodeficiencies. The results should also provide insights into the mechanisms involved in mucosal immunity to other intracellular pathogens.